Hi Nestor,
I think the reason for Arg side chain to curl up is because I refined
ligand and Arg side chain with occupancy refinement, and the Arg moved
away from the density, most likely due to 'repulsion' from ligand.
The other question is: Is it possible that the H-bonds stay very
close? As I tried to 'real space refine' in coot, and the Arg side
chain flipped away from the density.
Thanks for the suggestion.
Best
HK
Quoting Nestor Concha <nestor.o.con...@gsk.com>:
Hi Tam,
The density looks very strong and therefore I'm going to guess that
the Arg guanidimium stays in contact/interacts with the ligand and
with the phosphate/sulfate next to it. Perhaps it is one of those
close interactions with shorter H-bonds that usual given the
arrangement of ligand-phosphate/sulfate-Arg. I'd try to find a
rotamer for the Arg that leaves the interactions intact rather than
refine occupancies. Seems that the Arg side chain is curled up ????
Nestor
-----Original Message-----
From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> On Behalf Of Heng-Keat Tam
Sent: Tuesday, November 5, 2019 10:55 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Occupancy refinement of overlapping electron
density of a residue and ligand
EXTERNAL
Dear Rob,
I would like to model the alternative position for the side chain of
R120 but I don't really know whether the alternative conformation
exist as shown in the attached figure - density without the ligand and
R120 overlaid with the refined structures of modeled ligand and R120.
The ligand was modeled in two different conformations. From the
density, it seems to me that the density is connected or overlapped.
It should not be a post-translation modification as it is well-known
that there is no post-translation modification for this protein.
Furthermore, the crystal was obtained by co-crystallization of
protein with the ligand itself. The density seems to be the expected
ligand.
Thanks for the advice.
Best regards
HK
Quoting Robert Nicholls <nicho...@mrc-lmb.cam.ac.uk>:
Dear HK,
No that's not quite correct - 'occupancy group alts complete' means
that both R120 and the ligand are constrained so that their
occupancies sum to unity. In contrast, 'occupancy group alts
incomplete' means that the occupancies of R120 and the ligand will not
be constrained to sum to unity (but the sum of their occupancies must
be less than one). In both cases, R120 and the ligand will "see" each
other in a certain sense. But, because they are assigned to different
groups, it is assumed that they are present in different parts of the
crystal. This means that they can overlap.
Assuming that the ligand is in the correct conformation, I suspect the
source of your problem is that you are modelling the side chain of
R120 as only one conformation. And I would also include the other
atoms in the side chain - CB and CG.
If you are modelling the sidechain of R120 with partial occupancies,
then you should model those side chain atoms in two alternative
positions (i.e. representing the portions of the crystal that do/don't
have the ligand bound). This will help to ensure that your model makes
physical sense. So the ligand plus the alt of R120 in the portion of
the crystal that contains the ligand would be assigned to one
occupancy group, and the alt of R120 in the portion of the crystal
that does not contain the ligand would be assigned to the second
group. In this case it would be appropriate to specify 'occupancy
group alts complete', because the occupancies for the two alternative
conformers of R120 should sum to unity. Although no doubt the
estimation of the occupancies would be dominated by the ligand in this
case.
Be sure to check your B-factors after occupancy refinement to make
sure the whole picture makes sense. Assuming your current model is
essentially correct, from visual inspection it looks like R120 will
have low occupancy and low B-factors when the ligand is not present
(or at least B-factors consistent with the environment), but will have
high occupancy and high B-factors when the ligand is present.
On another note, have you considered whether that part of the ligand
could be modelled in a slightly different conformation, or whether
there could be a post-translation modification?
I hope that helps,
Rob
Dr Rob Nicholls
Senior Investigator Scientist
MRC Laboratory of Molecular Biology
Francis Crick Avenue
Cambridge Biomedical Campus
Cambridge CB2 0QH
On 5 Nov 2019, at 13:36, HK <t...@em.uni-frankfurt.de> wrote:
Dear all,
I have problem with occupancy refinement by Refmac5 for an
overlapping electron density of part of residue (an arginine) and
part of ligand (tetracyclic compound) (attached figures in Dropbox
with a link as shown below).
https://www.dropbox.com/sh/ppmfp5dnpy1b9e9/AAAV79bOzPHQUrVYp9loMwyha?
dl=0
I refined part of the side chain of residue 120 and ligand (chain J
residue 1105) with Refmac keyword as shown below. Unfortunately, the
side chain of arginine moves away from the density but the ligand
stays in the density. As far as I understood, occupancy refinement
with keyword 'occupancy group alts complete' means both
R120 and the ligand do not meet each other. Did I miss something
from the occupancy refinement keyword?
occupancy group id 1 chain A residue 120 atom NE occupancy group id 1
chain A residue 120 atom CZ occupancy group id 1 chain A residue 120
atom NH2 occupancy group id 1 chain A residue 120 atom NH1 occupancy
group id 1 chain A residue 120 atom CD occupancy group id 2 chain J
residue 1105 occupancy group alts complete 1 2 occupancy refine
Thank you for the advice.
Best regards
HK
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